In biology, angiogenesis refers to a process of generation of new blood vessels by budding or dividing from the existing blood vessels (capillaries, small arteries and veins) in the body. Angiogenesis is beneficial and essential for maintaining many normal physiological processes, such as embryonic development, wound healing, and repair. On the other hand, excessive blood vessel proliferation or angiogenesis is also associated with pathological processes, such as tumor growth, metastasis and inflammation. The key reason for the in vivo proliferation of the blood vessels is due to the ability of endothelial cells to divide and proliferate incisively and to insert to the existing vascular wall. Vascular endothelial growth factor (VEGF) is the most important and most potent angiogenic factor responsible for vascular endothelial cell growth. The importance of VEGF in angiogenesis has been well demonstrated by studies in VEGF knock-out mice: mouse embryos carrying one copy of VEGF gene, while the other one was knocked-out, would die at 11 to 12 days of development due to a decrease and abnormality in vascular angiogenesis (Carmeliet P et. al., Nature 1996, 380:435; Ferrara N et. al., Nature 196, 380:439).
Overexpression of VEGF has been observed in a variety of malignancies, such as colorectal, stomach, ovarian, breast cancers, hepatocelluar carcinoma and multiple myeloma; the level of VEGF expression is highly correlated with tumor growth, relapse and metastasis (Dvorak H F et. al: J Exp Med 1991; 174:1275-8; Brown L F et. al., Cancer Res 1993; 53:4727-35; Weidner N, Semple J P, Welch W R and Folkman J: N Engl J Med 1991; 324:1-8.4-5). In recent years, accumulating data from a series of animal experiments have shown that blocking angiogenesis by inhibiting the interaction of VEGF and VEGF-R, via gene manipulation or administration of drugs, leads to tumor ischemia and necrosis, which in turn, results in inhibition of tumor growth, metastasis and ultimately, prolongation of overall survival. Therefore, drug development targeting VEGF mediated angiogenesis has become a hot area of research worldwide.
Currently there are two major approaches to the anti-angiogenesis drug development targeting VEGF and VEGF-R pathway.
The first approach involves an inhibitor that antagonizes the tyrosine kinase located in the intracellular domain of the VEGF receptor (VEGF-R). Such antagonistic inhibitors are generally small-molecule chemical drugs, the prototypic drugs include Sutent (Sunitinib), which was developed and made available on the market in 2006 by Pfizer, and Nexavar (Sorafenib), developed and marketed by Bayer (Germany) and Onyx Pharmaceuticals.
The second approach involves large protein molecules, either an antibody or a fusion protein employing the Fc receptor of an antibody, which can directly block the binding of VEGF to its receptor (VEGF-R). One drug developed via this approach is Avastin (Bevacizumab), a humanized anti-VEGF monoclonal antibody drug which was developed and produced by Roche/Genentech and obtained FDA approval in February 2004. Avastin is so far the only anti-VEGF monoclonal antibody available on the global market. Avastin, by highly specific binding to VEGF, prevents VEGF from binding to VEGF-R, and thus blocks angiogenesis and inhibits tumor proliferation (Presta L G et. al., Cancer Res, 1997, 57: 4593; Hurwitz H et. al., N Engl J Med, 2004; 350:2335). Avastin has currently been approved by the FDA to be used for treatment of metastatic colorectal cancer (mCRC), advanced non-squamous non-small cell lung cancer (NSCLC), glioblastoma, metastatic renal cell carcinoma (mRCC), and various other solid tumors. Avastin also received approval from China SFDA in February 2010 for the treatment of colon cancer.
The precursor of Avastin can be traced back to a mouse monoclonal antibody, A4.6.1. The origin of this antibody, the hybridoma cell lines secreting it, and its use are described in the following patents: U.S. Pat. No. 6,582,959 (inventor: Kim, Kyung Jin; patent date: Jun. 24, 2003; patent title: Antibodies to Vascular Endothelial Growth Factor); and U.S. Pat. No. 7,227,004 (Inventor: Kim, Kyung Jin; patent date: Jun. 5, 2007; patent title: Antibodies to Vascular Endothelial Growth Factor). The amino acid sequence of this murine antibody and its humanized version, rhuMab-VEGF (i.e. Avastin) was published (Presta L G et. al., Cancer Res, 1997, 57: 4593). The method of preparation was disclosed in U.S. Pat. No. 6,054,297 (Inventor: Carter; Paul J. and Presta; Leonard G; patent application date: May 9, 1995; patent date: Apr. 25, 2000; patent title: Humanized Antibodies and Methods for Making Them).
However, this antibody still has the following shortcomings:
1) Similar to most other monoclonal antibodies, monoclonal antibody A4.6.1 (or Avastin) can only bind to a part of VEGF region or epitope, but cannot bind to other epitopes or cover other areas of VEGF antigen.
2) Previous animal experiments and recent clinic research have shown that administering A4.6.1 or Avastin alone was not able to neutralize VEGF entirely, or completely inhibiting VEGF mediated angiogenesis in vivo.
Therefore, it is important and necessary to develop new monoclonal antibodies or therapeutic agents which have the ability to bind to VEGF on new binding sites and same time can inhibit the binding of VEGF to VEGF-R.